1. Field of the Invention
The present invention relates to a gene encoding a polypeptide possessing lacto-N-biosidase activity, and to a method for producing a polypeptide possessing lacto-N-biosidase activity by the use of the gene.
2. Discussion of the Related Art
Considerable attention has been given in recent years to various physiological functions of the sugar chains of so called complex carbohydrates, such as glycoproteins and glycolipids existing on the surface of animal cells. A glycosidase with high specificity has become a very useful tool in the research of structures and biological activities of the sugar chains. Two types of sugar chains, type 1 structure (Gal .beta.1-3GlcNAc .beta.1-) and type 2 structure (Gal .beta.1-4GlcNAc .beta.1-) are frequently found in N- and O-linked sugar chains of glycoproteins, or in sugar chains of glycolipids. In the structural analyses of these sugar chains, lacto-N-biosidase is used as a very useful reagent, because lacto-N-biosidase specifically acts on the type 1 structure and is capable of releasing lacto-N-biose (Gal .beta.1-3GlcNAc) from the nonreducing terminus.
With respect to lacto-N-biosidase, those produced by Streptomyces strains are known Proceedings of the National Academy of Sciences of the USA, 89, 8512-8516 (1992); Journal of Biological Chemistry, 268, 18560-18566(1993)!.
A method for producing lacto-N-biosidase by culturing such lacto-N-biosidase producing Streptomyces bacteria requires tedious and time-consuming purification procedures because the bacteria also produce other enzymes such as protease, .beta.-N-acetylglucosaminidase and .alpha.-1,3/4-fucosidase. Also, in order to induce enzyme production, it is required to add L-fucose, swine gastric mucin, etc. Hence, there has been a need for a method producing lacto-N-biosidase at low cost and high purity.
A method for purification of lacto-N-biosidase from a culture of Streptomyces strains is known (Japanese Patent Laid-Open No. 6-153944). However, neither the amino acid sequence of lacto-N-biosidase nor the gene sequence encoding the enzyme has so far been elucidated, thereby impeding the production of lacto-N-biosidase by recombinant DNA technology.